The goal of this proposal is to elucidate the mechanisms by which the adjuvant GM-CSF enhances vaccine induced IgG and IgA responses against SIV. These studies will take advantage ofthe unique resources made available by this consortium and ofthe complementary and integrative expertise ofthe Amara and Pulendran groups, which evaluate novel Env immunogens for the induction of neutralizing Abs (NAbs) (Project 1 and Project 2). the Ahmed/Silvestri/Crotty group, which explores the regulation of Ab responses by T cells (Project 3). and the Cerutti group, which studies the regulation of B cells by innate immune cells (Project 4). B cells provide immune protection against HIV by producing NAbs to envelope (Env) spikes on the surface ofthe virus. However, eliciting robust and sustained NAb responses remains a major obstacle, because Env, the only relevant antigen for NAb induction, is characterized by sequence variation, limited antigenicity and scarce immunogenicity. An additional obstacle relates to the lack of strategies capable of effectively inducing NAbs both systemically and at mucosal sites of entry. Preliminary data from the Amara group show that GM-CSF enhances the avidity and frequency of vaccine-induced SIV-reactive IgG Abs produced in systemic lymphoid organs and elicits release of SIV-specific IgA in intestinal secretions. These effects correlate with increased protection against an intestinal challenge. In this proposal we hypothesize that GM-CSF mobilizes and activates a unique subset of splenic IL-21-producing NBH neutrophils equipped with B cell helper function. We contend that NBH cells enhance systemic IgG and intestinal IgA responses against SIV by inducing Ig heavy chain class switching, V(D)J gene somafic hypermutation and gut-homing receptors in splenic B cells, including marginal zone and memory B cells. Three aims are proposed. Aim 1 is to elucidate the mechanism by which GM-CSF induces IgG and IgA class switching in splenic B cells. Aim 2 is to dissect the mechanism by which GM-CSF induces intestinal homing of splenic IgA class-switched B cells. Aim 3 is to determine the mechanism by which GM-CSF improves the avidity of vaccine-induced systemic IgG and intestinal IgA responses against SIV..